The long-term objective is to understand the operation of metal-ion clusters and electron/proton transfer processes in biology. The specific goal is to understand the mechanism of photosynthetic water oxidation. Oxygenic photosynthesis provides the molecular oxygen and fixed carbon required to sustain all animal life. The proposed work will identify amino acid residues that coordinate the manganese and calcium ions at the catalytic site of water oxidation or whose protonation properties influence the reactivity of the manganese cluster. Identification of these residues will impose new constraints on models for the operation of the manganese cluster, and will enable such models to be placed in the context of protein structure. The information obtained will be applicable to other metal-ion clusters, and to the study of membrane proteins and biological electron and proton transfer processes in general. Electron and proton transfer reactions can be studied more easily and precisely in photosynthetic than in mitochondrial systems because both processes can be conveniently initiated with light. In oxygenic photosynthesis, Photosystem II (pSII) uses light to extract electrons from water and donate them into an electron transport chain that generates the chemical free energy and reducing equivalents required for carbon fixation. PSII photochemistry takes place in a heterodimer of two polypeptides known as D1 and D2. A cluster of four manganese ions accumulates four oxidizing equivalents in response to this photochemistry, then oxidizes two molecule of water by a process that requires calcium and releases molecular oxygen as a by-product. The proposed work will (l) determine if specific amino acid residues of the D1 polypeptide (e.g, Asp- 170, His-332, Glu-333, His-337, and Asp-342) ligate the Mn cluster, (2) characterize the role of specific amino acid residues of the D1 polypeptide in the binding of calcium (e.g., Asp-59, Asp-61, His-332, Glu- 333, His-337, and Asp-342), (3) further characterize the influence of particular residues of the D1 polypeptide on the operation of the intact Mn cluster (e.g., Asp61, His-92, and Glu-189), and (4) identify amino acid residues whose protonation properties influence the energetics and mechanism of water oxidation. Mutants with substitutions of the latter residues would contain Mn clusters that are significantly perturbed or are non-functional and that exhibit altered patterns of proton release. The required mutations have already been constructed by us in the cyanobacterium Synechocystis sp. PCC 6803.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM043496-08
Application #
2444746
Study Section
Molecular and Cellular Biophysics Study Section (BBCA)
Project Start
1989-12-01
Project End
1999-06-30
Budget Start
1997-07-01
Budget End
1998-06-30
Support Year
8
Fiscal Year
1997
Total Cost
Indirect Cost
Name
University of California Riverside
Department
Biochemistry
Type
Schools of Earth Sciences/Natur
DUNS #
City
Riverside
State
CA
Country
United States
Zip Code
92521
Debus, R J; Campbell, K A; Gregor, W et al. (2001) Does histidine 332 of the D1 polypeptide ligate the manganese cluster in photosystem II? An electron spin echo envelope modulation study. Biochemistry 40:3690-9
Debus, R J (2001) Amino acid residues that modulate the properties of tyrosine Y(Z) and the manganese cluster in the water oxidizing complex of photosystem II. Biochim Biophys Acta 1503:164-86
Chu, H A; Debus, R J; Babcock, G T (2001) D1-Asp170 is structurally coupled to the oxygen evolving complex in photosystem II as revealed by light-induced Fourier transform infrared difference spectroscopy. Biochemistry 40:2312-6
Faller, P; Debus, R J; Brettel, K et al. (2001) Rapid formation of the stable tyrosyl radical in photosystem II. Proc Natl Acad Sci U S A 98:14368-73
Debus, R J; Campbell, K A; Pham, D P et al. (2000) Glutamate 189 of the D1 polypeptide modulates the magnetic and redox properties of the manganese cluster and tyrosine Y(Z) in photosystem II. Biochemistry 39:6275-87
Debus, R J; Campbell, K A; Peloquin, J M et al. (2000) Histidine 332 of the D1 polypeptide modulates the magnetic and redox properties of the manganese cluster and tyrosine Y(Z) in photosystem II. Biochemistry 39:470-8
Debus, R J (2000) The polypeptides of photosystem II and their influence on manganotyrosyl-based oxygen evolution. Met Ions Biol Syst 37:657-711
Qian, M; Dao, L; Debus, R J et al. (1999) Impact of mutations within the putative Ca2+-binding lumenal interhelical a-b loop of the photosystem II D1 protein on the kinetics of photoactivation and H2O-oxidation in Synechocystis sp. PCC6803. Biochemistry 38:6070-81
Hays, A M; Vassiliev, I R; Golbeck, J H et al. (1999) Role of D1-His190 in the proton-coupled oxidation of tyrosine YZ in manganese-depleted photosystem II. Biochemistry 38:11851-65
Hundelt, M; Hays, A M; Debus, R J et al. (1998) Oxygenic photosystem II: the mutation D1-D61N in Synechocystis sp. PCC 6803 retards S-state transitions without affecting electron transfer from YZ to P680+. Biochemistry 37:14450-6

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